This project seeks to examine specific excitation-contraction coupling mechanisms in bladder smooth muscle cells, and to determine whether these processes are associated with the contractile dysfunction known to occur with chronic obstruction of the urinary bladder. Particular focus will be on the molecular mechanisms underlying calcium-induced calcium release (CICR) in bladder smooth muscle cells, using laser scanning confocal microscopy and patch clamp methods. Calcium-induced calcium release, or the release of intracellular calcium ions by the influx of calcium across the cell membrane, has been shown to play an important role in excitation-contraction coupling in bladder myocytes. Evidence suggests that impairment of this process is associated with the adaptive changes that occur in bladder smooth muscle during outlet obstruction. We will determine the biophysical characteristics of CICR in normal urinary bladder myocytes and compare that with observations in myocytes from obstructed bladders. Knockout mice with targeted deletions of ryanodine receptors will be generated and the CICR process examined to determine the role of specific intracellular calcium channels and associated modulatory proteins in excitation-contraction coupling of bladder smooth muscle.